Arthropod-borne viruses (arboviruses) are maintained in nature in cycles involving vertebrate hosts and hematophagous arthropods. Arboviruses can cause devastating diseases in vertebrate hosts, yet establish life-long persistent infections in vectors. The most amazing of arbovirus-vector interactions is transovarial transmission (TOT). LaCrosse (LAC) virus replicates in Aedes triseriatus mosquitoes during critical periods of vector oogenesis and embryogenesis without adverse effects. Co-regulation of vector and virus gene expression apparently modulates viral virulence in critical life stages of the vector. Although modulation of virulence is necessary, the arbovirus must also elude components of the vector innate immune response, e.g., apoptosis and RNAi, in order to persist. LAC (Bunyaviridae) and West Nile (WN, Flaviviridae) viruses induce apoptosis in vertebrate cells, but not in vector cells. Exogenously induced RNA interference (RNAi) inhibits both dengue (DEN; Flaviviridae) and LAC virus infection of mosquito cells, but we do not know if natural infections induce RNAi. In these studies, we will elucidate the roles of apoptosis and RNAi in arboviral persistence. LAC and WN virus will be used in comparative studies to elucidate whether apoptosis and RNAi are determinants of arbovirus attenuation and persistence in vectors. Both viruses induce apoptosis in vertebrate cells. LACV does not induce apoptosis in vector cells, is absolutely remarkable in its ability to establish long-term persistent infections in vectors, and is very efficiently transmitted TOT. WNV does not induce apoptosis in vector cells, establishes persistent infections in adult vectors, but is not transmitted TOT. The underlying hypothesis for these studies is that the replicative strategies of these two viruses determines their different phenotypes in vectors. RNAi and apoptosis can be triggered by dsRNA molecules. Replication and transcription of bunyaviruses differs fundamentally from that of flaviviruses; the dsRNA replicative intermediates of the two families differ dramatically, which could result in bunyavirus avoidance of both dsRNA-induced apoptosis and RNAi in vectors. Modern molecular biological and genomic approaches will be used to define the role of apoptosis and RNAi in persistent arbovirus infections in invertebrate hosts. Special emphasis will be devoted to determining the role of RNAi and apoptosis in TOT of arboviruses, which serves both to maintain trans-seasonally and to amplify bunyaviruses (but not flaviviruses) in nature. These studies may reveal fundamental molecular processes that condition these biologically and epidemiologically important vector phenotypes.